Cryopreservation and hypothermic storage of lacrimal gland: towards enabling delivery of regenerative medicine therapies for treatment of dry eye syndrome

J Tissue Eng Regen Med. 2017 Dec;11(12):3373-3384. doi: 10.1002/term.2251. Epub 2016 Dec 14.

Abstract

Severe dry eye syndrome (DES) can cause painful loss of vision and may result from lacrimal gland dysfunction. Current treatments are palliative, so a causative therapy is desirable. The ability to (cryo)preserve lacrimal gland tissue or epithelial cells would simplify this. Here, lacrimal gland tissue was cryopreserved in 10% dimethylsulphoxide in liquid nitrogen, or stored at 4 °C in culture medium for up to 7 days, and compared with fresh tissue using immunohistochemistry. Cultures were initiated from fresh and stored tissue, and cells characterised in P1 for proliferation (WST-1), colony-forming efficiency (CFE) and secretory capacity (immunocytochemistry and β-hexosaminidase activity assay). Tissue stored for > 3 days at 4 °C displayed grossly altered tissue architecture when compared with fresh tissue, decreased acinus density and increased caspase-3 activity. Cryopreserved tissue showed less obvious signs of damage without caspase-3 activation. Storage at 4 °C and cryopreservation delayed epithelial outgrowth compared with that from fresh tissue initially (p < 0.05) but, by day 9, all explants showed comparable outgrowth (~90%), except tissue stored at 4 °C for 3 or 7 days (p < 0.05 compared with fresh tissue). Epithelial cell yields per explant were similar from fresh and stored tissue, apart from tissue stored at 4 °C for 7 days (p < 0.01). In P1, epithelial cells from fresh and stored tissue were largely equivalent in terms of: proliferation; CFE (~21%); Rab3D, HexA and lysozyme expression; mucin production; and β-hexosaminidase activity. These data demonstrate that cryo(preservation) of lacrimal gland tissue and cells is possible, which may enable use of autologous cells in regenerative medicine approaches to treating DES. Copyright © 2016 John Wiley & Sons, Ltd.

Keywords: cryopreservation; dry eye syndrome; hypothermic storage; lacrimal gland; regeneration; tissue engineering.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3T3 Cells
  • Animals
  • Caspases / metabolism
  • Cell Proliferation
  • Cell Survival
  • Cold Temperature*
  • Colony-Forming Units Assay
  • Cryopreservation*
  • Dry Eye Syndromes / therapy*
  • Enzyme Activation
  • Epithelial Cells / cytology
  • Lacrimal Apparatus / transplantation*
  • Mice
  • Mucins / metabolism
  • Regenerative Medicine / methods*
  • Sus scrofa
  • Time Factors

Substances

  • Mucins
  • Caspases